Filling out the soft tissues is performed in plastic surgery to correct skin defects such as wrinkles, facial grooves and pitting. It can also increase the volume of particular areas such as deep scars, the lips and cheekbones, and better define the facial features and shape. These results are obtained by injecting fillers into the superficial or deep dermis to swell the area to be treated, making it firmer. Besides filling the depression, the injection triggers a phase of biostimulation of the skin cells, so that the skin itself looks healthier, firmer and rosier.
The substances used are called fillers and they are many and various. They can be substantially differentiated into three different types:                bioresorbable fillers; biocompatible substances that are subject to gradual and ultimately complete resorption by the organism. The most commonly used are collagen (Zyderm®, Zyplast®) and hyaluronic acid (Hylaform®, Ial System®, Restylane®) which give good results, especially in the correction of mild to medium defects, which are the most commonly treated. These materials are however limited because they may prove allergenic (especially collagen), in the presence of contaminating biological material (such as viruses or protein residues) due to the extraction process, and, more importantly, they require frequent administration in order to maintain their effect. Indeed, these are substances, hyaluronic acid in particular, that are rapidly degraded both by the enzymes and the free radicals that are physiologically present in the dermis. The resulting turgor can only be maintained by frequent booster injections of the product, with a consequent increase in the risk of side effects and discomfort to the patient;        Semi-permanent fillers, that last longer once they have been implanted in the tissues, as they are constituted by a bioresorbable matrix which incorporates particles such as polymethacrylate or acrylic hydrogel or dextran (among the commercial products of this kind are Artecoll®, Dermalive® and Reviderm® Intra). After resorption of the matrices, the non-biodegradable particles do maintain a certain degree of turgor but they may also cause inflammatory phenomena and marked allergic reactions;        permanent fillers, that are not resorbed by the organism. The products are based on hydrogels of polyacrylamide, Gore-Tex® or other completely synthetic materials which, after implantation, become progressively surrounded by a capsule of connective tissue that fixes them firmly in place. If on the one hand this is an advantage, because it renders the implant permanent, on the other it makes it difficult, but theoretically not impossible, to alter the effect or remove the implant if the desired effect is not achieved. Implanting permanent fillers is a surgical procedure, so the risks and benefits must be weighed up, creating a further limitation.        
The choice of filler is based on a series of parameters such as the desired effect and its duration, biocompatibility, painfulness, the possible need for allergy tests beforehand and the cost. In the field of bioresorbable fillers, one of the key factors when choosing is certainly the duration of the implant. Indeed, it is essential to choose a product that not only has all the aforesaid properties but also stays at the injection site for a long time, so as to reduce the number of administrations necessary to maintain the effect. This translates into a lesser risk of side effects due to the injection procedure (e.g. swelling, intumescence, burning) and consequently less discomfort for the patient. The limitations of the current state of the art have been overcome by the present invention, which describes and claims a bioresorbable filler based on hyaluronic acid and/or the derivatives thereof, structured with/in phospholipid liposomes that increase their residence time and improve their overall performance.
Hyaluronic acid (HA) is a well-known molecule: it is a heteropolysaccharide constituted by D-glucuronic acid and N-acetyl-glucosamine, and is present in practically every compartment of our organism. HA plays numerous physiological roles, ranging from mechanical support for the cells of many tissues to joint lubrication, the modulation of many biological and physiological processes (including cell proliferation, migration and differentiation, mediated by the interaction with its membrane receptor, CD44). HA's protective effect against the degeneration of cartilage that has been damaged by disease or trauma is well known. In such situations there is a strong concentration of pro-inflammatory cytokines in the joint cavity, especially interleukine-1 (IL-1), that promote cartilage disintegration and inhibit chondrocyte proliferation (van Beuningen H. M. et al., Arthritis Rheum, 1991, 34:606-615). Various scientific experiments have demonstrated that hyaluronic acid is able to oppose the action of IL-1, drastically reducing its negative effects and then exercising a reparatory effect on the cartilage tissue in the joint into which it has been injected. (Stove J. et al., J Orthop Res, 2002, 20:551-555). In the joints, the hyaluronic acid content in the synovial fluid acts as a viscous lubricant during slow movement, while during brisk movement its elastic properties absorb any trauma or microtrauma that may affect the joint. In pathological situations, both the concentration and mean molecular weight of HA (Balazs E A. et al., J Rheumatol Suppl, 1993, 12:75-82; Belcher C. et al., Annals of the Rheumatic Disease, 1997, 56:299-307) decrease considerably, altering the physiological features of the synovial fluid.
Its tissue-hydrating and wound-healing properties are also widely known and have long been put to use in medications for the treatment of wounds, ulcers and skin lesions of various origin (e.g., Balasz A. et al., Cosmetics & Toiletries, 1984, 5:8-17).
Numerous chemical modifications that can be performed on the HA molecule are also known to the state of the art, that is:    salification with organic and/or inorganic bases (EP 138572 B1);    esterification of HA with alcohols of the aliphatic, araliphatic, cycloaliphatic, aromatic, cyclic and heterocyclic series (HYAFF®), with a percentage of esterification that may vary according to the type and length of the alcohol that is used (EP 216453 B1);    amidation of HA with amines of the aliphatic, araliphatic, cycloaliphatic, aromatic, cyclic and heterocyclic series (HYADD™), with a percentage of amidation ranging between 0.1 and 50% (EP 1095064 B1);    O-sulphatation of HA to the 4th degree of sulphatation (EP 702699 B1);    Inner esterification of HA with a percentage of esterification not exceeding 20% (ACP®; EP 341745 B1); deacetylation of HA: the N-acetyl-glucosamine fraction is deacetylated, preferably to a percentage of between 0.1 and 30% (EP 1313772 B1);    percarboxylation of HA achieved by oxidising the primary hydroxyl of the N-acetyl-glucosamine fraction to a degree of percarboxylation of between 0.1 and 100% (HYOXX™; patent application EP 1339753).
The polymers obtained by these processes maintain the characteristics of biodegradability, biocompatibility, and easy handling and use of the starting polysaccharide, but they give a better mechanical performance.
The hyaluronic acid used in the present invention may derive from any source. For example, it may be extracted from rooster combs (EP 138572 B1) or obtained by fermentation (EP 716688 B1) or by technological means, and its molecular weight may range between 50,000 and 3,000,000 Da.
The type of technical solution described and claimed in the present invention is, however, absolutely innovative, and the fillers of HA and/or the derivatives thereof therefore remain at the application site for a long time, significantly reducing the need for frequent administrations while maintaining the characteristics of biocompatibility, safety and easy handling and use of the starting polysaccharide. This characteristic is achieved by structuring the hyaluronic acid and/or the derivatives thereof with/in phospholipid liposomes, as illustrated hereafter. Liposomes are hollow microspheres of varying size, ranging between 50 nm and 1000 nm, formed by one or more double lipid layers that enclose a hydrophilic core. This structure can be achieved thanks to the special nature of phospholipids that have a hydrophobic tail and hydrophilic head; in an aqueous medium the hydrophobic tails attract one another while the hydrophilic heads tend to face water. The result is double lipid layers that close to form small vesicles inside which there is a variously hydrophilic environment. Liposomes were first described in 1965 (Standish MM et al., J Mol Biol, 1965, 13:238-252) and have been researched as carriers for drugs and/or active ingredients (e.g., Liposomes as drug carriers, Gregoriadis G. editor, New York: John Wiley & Sons, 1985: 3-18; Banerjee R., J Biomater Appl, 2001, 16:3-21). They are normally classified on the basis of their size and the number of double lipid layers. Generally speaking, as described, for example, by Callow R A et al. (Cryobiology, 1985:251-267), reference is made to    multilamellar vesicles: they have an onion-like structure wherein a number of double lipid layers are interspersed with hydrophilic layers;    unilamellar vesicles, large (diameter of over 1 μm) and small (diameter of less than 1 μm): they are formed by one single double lipid layer and enclose a strongly hydrophilic nucleus;    oligolamellar vesicles, constituted by several double lipid layers that enclose a markedly hydrophobic environment.
Further classifications are possible on the basis of numerous processes by which liposomes can be obtained and which are well known to the expert in the field. Combinations of HA and phospholipids have already been described both as simple physical mixtures (WO 91/12026) and as proper chemical associations (EP 581282 B1) intended for use as antirheumatic drugs for intra-articular use, for which the lubricating properties of both liposomes and the polysaccharides in question are claimed. Also known is patent application EP 1406571 that describes and claims the use of glycosaminoglycans encapsulated in phospholipid liposomes for the intra-articular treatment of osteoarthrosis.
The Applicant intends to demonstrate hereafter that the present invention differs substantially from those already known in the type of polysaccharide used and also in the way in which it is structured.